1. Field of the Invention
This invention pertains to an assay for the detection of levels of neutralizing antibodies effective against Infectious Bursal Disease Virus (IBDV) in poultry, specifically chickens. Particularly, a competitive ELISA is employed, using a labeled monoclonal antibody specific to all strains and serotypes of IBDV to determine a base line for the amount of this labeled antibody which will bind to a given amount of antigen (IBDV). The labeled antibody is admixed with the antigen and serum from the poultry, the reduced amount of binding between the labeled antibody and the antigen providing a reliable indication as to the amount of effective neutralizing antibody in the serum employed.
2. Background of the Prior Art
Infectious Bursal Disease Virus (IBDV) has been recognized as one of the principal economic drains on the poultry industry, responsible, alone, for the loss of extraordinarily high numbers of poultry individuals, and millions of dollars annually. In the disclosure of U.S. Patent Application 061,083, filed June 12, 1987, by Snyder et al, this problem, and the discovery of a monoclonal antibody specific to all known strains and serotypes of IBDV is disclosed in detail. Use of that monoclonal antibody (MCA) identified as R63 and expressed by the cell line deposited at the ATCC and bearing the Accession Number HB9490 and otherwise available from continuous viable cell lines maintained on the grounds of the University of Maryland, College Park campus, for conferring effective immunity against IBDV on new-born chicks is also disclosed in that application.
One problem widely encountered in the poultry industry in treating and preventing IBDV, whether it be through the use of MCA R63 or some other method, is identification of the level of antibody effective in neutralizing IBDV present in the serum of poultry individuals, and poultry flocks. A determination of the antibody level is necessary to determine the need for further immunization treatment, and the amount of treatment necessary, if appropriate.
Experience throughout the industry, and that of the inventor, indicates that the current method for determining antibody titer levels, an indirect-ELISA, does not give reliable results, the values obtained through this indirect assay and virus neutralization studies being, frequently, quite dissimilar.
It has been discovered that this lack of reproducability and reliability in results obtained through the currently marketed indirect-ELISA may be due to two significant factors associated with that assay. The current available assay measures not only antibodies effective in neutralizing IBDV (i.e., protective antibodies) but measures significant amounts of other antibodies which bind to the IBDV at non-neutralizing antigenic sites, and thus, gives a falsely high reading of antibody titer with respect to neutralization capacity of the serum. It should be noted that in the identified parent application of Snyder et al, it is disclosed that of the nine monoclonal antibodies developed in the study leading to the invention claimed therein, only two had any neutralizing effectiveness. Therefore, there are clearly non-neutralizing antigenic sites on the various strains of IBDV.
Currently available IBDV ELISA antibody kits also employ IBDV antigens that contain non-virus-related protein antigens which can be detected by probing with antisera monospecific to the suspect contaminants (e.g., fetal bovine serum bacterial antigens, fetal antigens contained in the serum, etc.). Antibodies which bind to these extraneous antigens are frequently present in the serum of vaccinated chickens and other poultry by virtue of vaccination with crude antigen preparations, and the binding of these antibodies is also measured falsely as anti-IBDV antibody in this indirect ELISA. Thus, currently available methods do not give reliable indications of the level of neutralizing, protective type IBDV antibody present in the serum of tested individuals.